Characterization of Homogeneous and Heterogeneous Amyloid-β42 Oligomer Preparations with Biochemical Methods and Infrared Spectroscopy Reveals a Correlation between Infrared Spectrum and Oligomer Size.

Faraz Vosough,Andreas Barth

doi:10.1021/acschemneuro.0c00642

Abstract

Soluble oligomers of the amyloid-β(1-42) (Aβ42) peptide, widely considered to be among the relevant neurotoxic species involved in Alzheimer’s disease, were characterized with a combination of biochemical and biophysical methods. Homogeneous and stable Aβ42 oligomers were prepared by treating monomeric solutions of the peptide with detergents. The prepared oligomeric solutions were analyzed with blue native and sodium dodecyl sulfate polyacrylamide gel electrophoresis, as well as with infrared (IR) spectroscopy. The IR spectra indicated a well-defined β-sheet structure of the prepared oligomers. We also found a relationship between the size/molecular weight of the Aβ42 oligomers and their IR spectra: The position of the main amide I′ band of the peptide backbone correlated with oligomer size, with larger oligomers being associated with lower wavenumbers. This relationship explained the time-dependent band shift observed in time-resolved IR studies of Aβ42 aggregation in the absence of detergents, during which the oligomer size increased. In addition, the bandwidth of the main IR band in the amide I′ region was found to become narrower with time in our time-resolved aggregation experiments, indicating a more homogeneous absorption of the β-sheets of the oligomers after several hours of aggregation. This is predominantly due to the consumption of smaller oligomers in the aggregation process.

Highlights

Pathological changes or behavioral symptoms associated with Alzheimer’s disease (AD) have been found to correlate with levels of soluble amyloid-β peptide (Aβ) in the brain or the cerebrospinal fluid.[1−4] As such, since about two decades ago, soluble oligomeric species have been mentioned as a major source of toxicity in AD, as well as in several other neurodegenerative diseases.[5]
Chemical crosslinking methods can be used to overcome this obstacle;[37] recent studies have revealed that even when combined with cross-linking methods, sodium dodecyl sulfate (SDS)-PAGE may be misleading in size estimation of Aβ oligomers.[38]
The band position correlates with oligomer size/molecular weight, because it depends on the twist and strand number of β-sheets, when the sheets are small

Summary

■ INTRODUCTION

Pathological changes or behavioral symptoms associated with Alzheimer’s disease (AD) have been found to correlate with levels of soluble amyloid-β peptide (Aβ) in the brain or the cerebrospinal fluid.[1−4] As such, since about two decades ago, soluble oligomeric species have been mentioned as a major source of toxicity in AD, as well as in several other neurodegenerative diseases.[5]. Aβ42 oligomers formed in absence of detergent are resistant to high SDS concentrations (2% SDS, as in the SDS-PAGE sample buffer) and appear largely as a smear between 50 and 250 kDa on the blot (lanes 2 and 3 for recombinant and synthetic peptide, respectively), which is similar to the molecular weight range in which a smear is observed with SDS-PAGE (Figure 5). A similar band shift between smaller and larger SDSstabilized oligomers has been observed in a preliminary report of this study using a simpler preparation for the monomeric peptide.[65] The band position of the globulomer preparation shifts to lower wavenumber with time (1626.8 cm−1 after 5 days), due to a slow formation of larger oligomers as observed previously with gel electrophoresis.[19] Oligomeric solutions prepared in the absence of any detergents are heterogeneous mixtures, which contain larger structures (Figure 6, lanes 2 and 3) and generate the main band at much lower wavenumbers, around 1621.9−1622.9 cm−1.

■ CONCLUSIONS

■ METHODS

■ ACKNOWLEDGMENTS

■ REFERENCES